This invention relates to nucleic acid and amino acid sequences of polypeptide subunits of NADH dehydrogenase and to the use of these sequences in the diagnosis, prevention, and treatment of cancer, myopathies, neurodegenerative diseases, immune system disorders, and diseases and disorders of the sympathetic nervous system.
NADH dehydrogenase (NADH:ubiquinone oxidoreductase, NADH-D) is the first multienzyme complex (Complex I) in a chain of three complexes that make up the mitochondrial electron transport chain. The mitochondrial electron transport chain is responsible for the transport of electrons from NADH to oxygen and the coupling of this oxidation to the synthesis of ATP (oxidative phosphorylation) which provides the energy source for driving a cell""s many energy-requiring reactions. NADH-D accomplishes the first step in this process by accepting electrons from NADH and passing them through a flavin molecule to ubiquinone, which transfers the electrons to the second enzyme complex in the chain.
NADH-D and the other members of the electron transport chain are located in the mitochondrial membrane. NADH-D is the largest of the three complexes with an estimated mass of 800 kDa comprising some 40 polypeptides of widely varying size and composition. The polypeptide composition of NADH-D in a variety of mammalian species including rat, rabbit, cow, and man is very similar (Cleeter, M. W. J. and Ragan, C. I. (1985) Biochem. J. 230: 739-46). The best characterized NADH-D is from bovine heart mitochondria and is composed of 41 polypeptide chains (Walker, J. E. et al. (1992) J. Mol. Biol. 226: 1051-72). Seven of these polypeptides are encoded by mitochondrial DNA while the remaining 34 are nuclear gene products that are imported into the mitochondria. These imported polypeptides are characterized by various N-terminal peptide sequences or modified N-terminal amino acids (myristoylation or acetylation) that target them to the mitochondria and are then cleaved from the mature protein. The 24-, 51-, and 75-kDa subunits have been identified as being catalytically important in electron transport, with the 51-kDa subunit forming part of the NADH binding site and containing the flavin moiety that is the initial electron acceptor. The location of other functionally important groups, such as the electron-carrying iron sulfate centers, remains to be determined. Many of the smaller subunits ( less than 30 kDa) may play a purely structural role in the complex.
Defects and altered expression of NADH-D are associated with a variety of disease conditions in man, incluing; neurodegenerative diseases, myopathies, and cancer. In addition, NADH-D reduction of the quinone moiety in chemotherapeutic agents such as doxorubicin is believed to contribute to the antitumor activity and/or mutagenicity of these drugs.
The discovery of polynucleotides encoding NADH dehydrogenase, and the molecules themselves, provides a means to investigate the control of cellular respiration under normal and disease conditions. Such molecules related to NADH dehydrogenase satisfy a need in the art by providing new diagnostic or therapeutic compositions useful in diagnosing and treating cancers, myopathies, neurodegenerative diseases, and diseases and disorders of the sympathetic nervous system.
The present invention features four NADH dehydrogenase subunits, designated individually as NDS-1, NDS-2, NDS-3 and NDS-4 and collectively as NDS, and characterized as having similarity to NADH dehydrogenase.
Accordingly, the invention features substantially purified NDS proteins NDS-1, NDS-2, NDS-3, and NDS-4 having the amino acid sequences shown in SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5 and SEQ ID NO:7, respectively.
One aspect of the invention features isolated and substantially purified polynucleotides that encode NDS proteins. In a particular aspect, the polynucleotides are the nucleotide sequences of SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:6, or SEQ ID NO:8 respectively.
The invention also features a polynucleotide sequence comprising the complement of SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8 or variants thereof. In addition, the invention features polynucleotide sequences which hybridize under stringent conditions to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:6, or SEQ ID NO:8.
The invention additionally features nucleic acid sequences encoding polypeptides, oligonucleotides, peptide nucleic acids (PNA), fragments, portions or antisense molecules thereof, and expression vectors and host cells comprising polynucleotides that encode NDS. The present invention also features antibodies which bind specifically to NDS, and pharmaceutical compositions comprising substantially purified NDS. The invention also features the use of agonists and antagonists of NDS.